Skip to main content
Top
Published in: Fluids and Barriers of the CNS 1/2013

Open Access 01-12-2013 | Short paper

Cytokine-induced changes in the gene expression profile of a human cerebral microvascular endothelial cell-line, hCMEC/D3

Authors: Miguel Alejandro Lopez-Ramirez, David Kingsley Male, Chunfang Wang, Basil Sharrack, Dongsheng Wu, Ignacio Andres Romero

Published in: Fluids and Barriers of the CNS | Issue 1/2013

Login to get access

Abstract

Background

The human cerebral microvascular endothelial cell line, hCMEC/D3, has been used extensively to model the blood–brain barrier (BBB) in vitro. Recently, we reported that cytokine-treatment induced loss of brain endothelial barrier properties. In this study, we further determined the gene expression pattern of hCMEC/D3 cells in response to activation with TNFα and IFNγ.

Findings

Using a microarray approach, we observed that expression of genes involved in the control of barrier permeability, including inter-brain endothelial junctions (e.g. claudin-5, MARVELD-2), integrin-focal adhesions complexes (e.g. integrin β1, ELMO-1) and transporter systems (e.g. ABCB1, SLC2A1), are altered by pro-inflammatory cytokines.

Conclusions

Our study shows that previously-described cytokine-induced changes in the pattern of gene expression of endothelium are reproduced in hCMEC/D3 cells, suggesting that this model is suitable to study inflammation at the BBB, while at the same time it has provided insights into novel key molecular processes that are altered in brain endothelium during neuroinflammation, such as modulation of cell-to-matrix contacts.
Appendix
Available only for authorised users
Literature
1.
go back to reference Coureuil M, Mikaty G, Miller F, Lecuyer H, Bernard C, Bourdoulous S, Dumenil G, Mege RM, Weksler BB, Romero IA, Couraud PO, Nassif X: Meningococcal type IV pili recruit the polarity complex to cross the brain endothelium. Science. 2009, 325: 83-87. 10.1126/science.1173196.PubMedCrossRef Coureuil M, Mikaty G, Miller F, Lecuyer H, Bernard C, Bourdoulous S, Dumenil G, Mege RM, Weksler BB, Romero IA, Couraud PO, Nassif X: Meningococcal type IV pili recruit the polarity complex to cross the brain endothelium. Science. 2009, 325: 83-87. 10.1126/science.1173196.PubMedCrossRef
2.
go back to reference Fletcher NF, Wilson GK, Murray J, Hu K, Lewis A, Reynolds GM, Stamataki Z, Meredith LW, Rowe IA, Luo G, Lopez-Ramirez MA, Baumert TF, Weksler B, Couraud PO, Kim KS, Romero IA, Jopling C, Morgello S, Balfe P, McKeating JA: Hepatitis C virus infects the endothelial cells of the blood–brain barrier. Gastroenterology. 2012, 142: 634-643. 10.1053/j.gastro.2011.11.028.PubMedCentralPubMedCrossRef Fletcher NF, Wilson GK, Murray J, Hu K, Lewis A, Reynolds GM, Stamataki Z, Meredith LW, Rowe IA, Luo G, Lopez-Ramirez MA, Baumert TF, Weksler B, Couraud PO, Kim KS, Romero IA, Jopling C, Morgello S, Balfe P, McKeating JA: Hepatitis C virus infects the endothelial cells of the blood–brain barrier. Gastroenterology. 2012, 142: 634-643. 10.1053/j.gastro.2011.11.028.PubMedCentralPubMedCrossRef
3.
go back to reference Bahbouhi B, Berthelot L, Pettre S, Michel L, Wiertlewski S, Weksler B, Romero IA, Miller F, Couraud PO, Brouard S, Laplaud DA, Soulillou JP: Peripheral blood CD4+ T lymphocytes from multiple sclerosis patients are characterized by higher PSGL-1 expression and transmigration capacity across a human blood–brain barrier-derived endothelial cell line. J Leukoc Biol. 2009, 86: 1049-1063. 10.1189/jlb.1008666.PubMedCrossRef Bahbouhi B, Berthelot L, Pettre S, Michel L, Wiertlewski S, Weksler B, Romero IA, Miller F, Couraud PO, Brouard S, Laplaud DA, Soulillou JP: Peripheral blood CD4+ T lymphocytes from multiple sclerosis patients are characterized by higher PSGL-1 expression and transmigration capacity across a human blood–brain barrier-derived endothelial cell line. J Leukoc Biol. 2009, 86: 1049-1063. 10.1189/jlb.1008666.PubMedCrossRef
4.
go back to reference Poller B, Drewe J, Krahenbuhl S, Huwyler J, Gutmann H: Regulation of BCRP (ABCG2) and P-glycoprotein (ABCB1) by cytokines in a model of the human blood–brain barrier. Cell Mol Neurobiol. 2010, 30: 63-70. 10.1007/s10571-009-9431-1.PubMedCrossRef Poller B, Drewe J, Krahenbuhl S, Huwyler J, Gutmann H: Regulation of BCRP (ABCG2) and P-glycoprotein (ABCB1) by cytokines in a model of the human blood–brain barrier. Cell Mol Neurobiol. 2010, 30: 63-70. 10.1007/s10571-009-9431-1.PubMedCrossRef
5.
go back to reference Tai LM, Reddy PS, Lopez-Ramirez MA, Davies HA, Male DK, Loughlin AJ, Romero IA: Polarized P-glycoprotein expression by the immortalised human brain endothelial cell line, hCMEC/D3, restricts apical-to-basolateral permeability to rhodamine 123. Brain Res. 2009, 1292: 14-24.PubMedCrossRef Tai LM, Reddy PS, Lopez-Ramirez MA, Davies HA, Male DK, Loughlin AJ, Romero IA: Polarized P-glycoprotein expression by the immortalised human brain endothelial cell line, hCMEC/D3, restricts apical-to-basolateral permeability to rhodamine 123. Brain Res. 2009, 1292: 14-24.PubMedCrossRef
6.
go back to reference Schreibelt G, Kooij G, Reijerkerk A, van Doorn R, Gringhuis SI, van der Pol S, Weksler BB, Romero IA, Couraud PO, Piontek J, Blasig IE, Dijkstra CD, Ronken E, de Vries HE: Reactive oxygen species alter brain endothelial tight junction dynamics via RhoA, PI3 kinase, and PKB signaling. FASEB J. 2007, 21: 3666-3676. 10.1096/fj.07-8329com.PubMedCrossRef Schreibelt G, Kooij G, Reijerkerk A, van Doorn R, Gringhuis SI, van der Pol S, Weksler BB, Romero IA, Couraud PO, Piontek J, Blasig IE, Dijkstra CD, Ronken E, de Vries HE: Reactive oxygen species alter brain endothelial tight junction dynamics via RhoA, PI3 kinase, and PKB signaling. FASEB J. 2007, 21: 3666-3676. 10.1096/fj.07-8329com.PubMedCrossRef
7.
go back to reference Afonso PV, Ozden S, Prevost MC, Schmitt C, Seilhean D, Weksler B, Couraud PO, Gessain A, Romero IA, Ceccaldi PE: Human blood–brain barrier disruption by retroviral-Infected lymphocytes: role of myosin light chain kinase in endothelial tight-junction disorganization. J Immunol. 2007, 179: 2576-2583.PubMedCrossRef Afonso PV, Ozden S, Prevost MC, Schmitt C, Seilhean D, Weksler B, Couraud PO, Gessain A, Romero IA, Ceccaldi PE: Human blood–brain barrier disruption by retroviral-Infected lymphocytes: role of myosin light chain kinase in endothelial tight-junction disorganization. J Immunol. 2007, 179: 2576-2583.PubMedCrossRef
8.
go back to reference Lopez-Ramirez MA, Fischer R, Torres-Badillo CC, Davies HA, Logan K, Pfizenmaier K, Male DK, Sharrack B, Romero IA: Role of caspases in cytokine-induced barrier breakdown in human brain endothelial cells. J Immunol. 2012, 189: 3130-3139. 10.4049/jimmunol.1103460.PubMedCrossRef Lopez-Ramirez MA, Fischer R, Torres-Badillo CC, Davies HA, Logan K, Pfizenmaier K, Male DK, Sharrack B, Romero IA: Role of caspases in cytokine-induced barrier breakdown in human brain endothelial cells. J Immunol. 2012, 189: 3130-3139. 10.4049/jimmunol.1103460.PubMedCrossRef
9.
go back to reference Forster C, Burek M, Romero IA, Weksler B, Couraud PO, Drenckhahn D: Differential effects of hydrocortisone and TNF{alpha} on tight junction proteins in an in vitro model of the human blood–brain barrier. J Physiol. 2008, 586: 1937-1949. 10.1113/jphysiol.2007.146852.PubMedCentralPubMedCrossRef Forster C, Burek M, Romero IA, Weksler B, Couraud PO, Drenckhahn D: Differential effects of hydrocortisone and TNF{alpha} on tight junction proteins in an in vitro model of the human blood–brain barrier. J Physiol. 2008, 586: 1937-1949. 10.1113/jphysiol.2007.146852.PubMedCentralPubMedCrossRef
10.
go back to reference Huang DW, Sherman BT, Lempicki RA: Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources. Nat Protocols. 2008, 4: 44-57. 10.1038/nprot.2008.211.CrossRef Huang DW, Sherman BT, Lempicki RA: Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources. Nat Protocols. 2008, 4: 44-57. 10.1038/nprot.2008.211.CrossRef
11.
go back to reference Viemann D, Goebeler M, Schmid S, Klimmek K, Sorg C, Ludwig S, Roth J: Transcriptional profiling of IKK2/NF-kappa B- and p38 MAP kinase-dependent gene expression in TNF-alpha-stimulated primary human endothelial cells. Blood. 2004, 103: 3365-3373. 10.1182/blood-2003-09-3296.PubMedCrossRef Viemann D, Goebeler M, Schmid S, Klimmek K, Sorg C, Ludwig S, Roth J: Transcriptional profiling of IKK2/NF-kappa B- and p38 MAP kinase-dependent gene expression in TNF-alpha-stimulated primary human endothelial cells. Blood. 2004, 103: 3365-3373. 10.1182/blood-2003-09-3296.PubMedCrossRef
12.
go back to reference Franzen B, Duvefelt K, Jonsson C, Engelhardt B, Ottervald J, Wickman M, Yang Y, Schuppe-Koistinen I: Gene and protein expression profiling of human cerebral endothelial cells activated with tumor necrosis factor-alpha. Brain Res Mol Brain Res. 2003, 115: 130-146. 10.1016/S0169-328X(03)00185-2.PubMedCrossRef Franzen B, Duvefelt K, Jonsson C, Engelhardt B, Ottervald J, Wickman M, Yang Y, Schuppe-Koistinen I: Gene and protein expression profiling of human cerebral endothelial cells activated with tumor necrosis factor-alpha. Brain Res Mol Brain Res. 2003, 115: 130-146. 10.1016/S0169-328X(03)00185-2.PubMedCrossRef
13.
go back to reference Kraft P, Benz PM, Austinat M, Brede ME, Schuh K, Walter U, Stoll G, Kleinschnitz C: deficiency of vasodilator-stimulated phosphoprotein (VASP) increases blood–brain-barrier damage and edema formation after ischemic stroke in mice. PloS one. 2010, 5: e15106-10.1371/journal.pone.0015106.PubMedCentralPubMedCrossRef Kraft P, Benz PM, Austinat M, Brede ME, Schuh K, Walter U, Stoll G, Kleinschnitz C: deficiency of vasodilator-stimulated phosphoprotein (VASP) increases blood–brain-barrier damage and edema formation after ischemic stroke in mice. PloS one. 2010, 5: e15106-10.1371/journal.pone.0015106.PubMedCentralPubMedCrossRef
14.
go back to reference Viemann D, Goebeler M, Schmid S, Nordhues U, Klimmek K, Sorg C, Roth J: TNF induces distinct gene expression programs in microvascular and macrovascular human endothelial cells. J Leukoc Biol. 2006, 80: 174-185. 10.1189/jlb.0905530.PubMedCrossRef Viemann D, Goebeler M, Schmid S, Nordhues U, Klimmek K, Sorg C, Roth J: TNF induces distinct gene expression programs in microvascular and macrovascular human endothelial cells. J Leukoc Biol. 2006, 80: 174-185. 10.1189/jlb.0905530.PubMedCrossRef
15.
go back to reference Nouvion AL, Oubaha M, Leblanc S, Davis EC, Jastrow H, Kammerer R, Breton V, Turbide C, Ergun S, Gratton JP, Beauchemin N: CEACAM1: a key regulator of vascular permeability. J Cell Sci. 2010, 123: 4221-4230. 10.1242/jcs.073635.PubMedCrossRef Nouvion AL, Oubaha M, Leblanc S, Davis EC, Jastrow H, Kammerer R, Breton V, Turbide C, Ergun S, Gratton JP, Beauchemin N: CEACAM1: a key regulator of vascular permeability. J Cell Sci. 2010, 123: 4221-4230. 10.1242/jcs.073635.PubMedCrossRef
16.
go back to reference Reijerkerk A, Lopez-Ramirez MA, van het Hof B, Joost AR, Drexhage WK, Kooij G, Joost BV, van der Pouw Kraan TCTM, Anton J, Van Z, Horrevoets AJ, Prat A, Romero IA, de Vries HE: microRNAs regulate human brain endothelial cell barrier function in inflammation: implications for multiple sclerosis. J Neurosci. 2013, 16: 6857-6863.CrossRef Reijerkerk A, Lopez-Ramirez MA, van het Hof B, Joost AR, Drexhage WK, Kooij G, Joost BV, van der Pouw Kraan TCTM, Anton J, Van Z, Horrevoets AJ, Prat A, Romero IA, de Vries HE: microRNAs regulate human brain endothelial cell barrier function in inflammation: implications for multiple sclerosis. J Neurosci. 2013, 16: 6857-6863.CrossRef
17.
go back to reference Lopez-Ramirez MA: The Open University, Department of Life, Health and Chemical Sciences. Investigating the role of inflammation-modulated microRNAs and signalling pathways in blood–brain barrier dysfunction. PhD Thesis. 2012, Lopez-Ramirez MA: The Open University, Department of Life, Health and Chemical Sciences. Investigating the role of inflammation-modulated microRNAs and signalling pathways in blood–brain barrier dysfunction. PhD Thesis. 2012,
18.
go back to reference McKenzie J, Ridley A: Roles of Rho/ROCK and MLCK in TNF-alpha-induced changes in endothelial morphology and permeability. J Cell Physiol. 2007, 213: 221-228. 10.1002/jcp.21114.PubMedCrossRef McKenzie J, Ridley A: Roles of Rho/ROCK and MLCK in TNF-alpha-induced changes in endothelial morphology and permeability. J Cell Physiol. 2007, 213: 221-228. 10.1002/jcp.21114.PubMedCrossRef
19.
go back to reference Daneman R, Zhou L, Kebede AA, Barres BA: Pericytes are required for blood–brain barrier integrity during embryogenesis. Nature. 2010, 468: 562-566. 10.1038/nature09513.PubMedCentralPubMedCrossRef Daneman R, Zhou L, Kebede AA, Barres BA: Pericytes are required for blood–brain barrier integrity during embryogenesis. Nature. 2010, 468: 562-566. 10.1038/nature09513.PubMedCentralPubMedCrossRef
20.
go back to reference Alexander JS, Elrod JW: Extracellular matrix, junctional integrity and matrix metalloproteinase interactions in endothelial permeability regulation. J Anat. 2002, 200: 561-574. 10.1046/j.1469-7580.2002.00057.x.PubMedCentralPubMedCrossRef Alexander JS, Elrod JW: Extracellular matrix, junctional integrity and matrix metalloproteinase interactions in endothelial permeability regulation. J Anat. 2002, 200: 561-574. 10.1046/j.1469-7580.2002.00057.x.PubMedCentralPubMedCrossRef
21.
go back to reference Schlegel N, Waschke J: Impaired integrin-mediated adhesion contributes to reduced barrier properties in VASP-deficient microvascular endothelium. J Cell Physiol. 2009, 220: 357-366. 10.1002/jcp.21772.PubMedCrossRef Schlegel N, Waschke J: Impaired integrin-mediated adhesion contributes to reduced barrier properties in VASP-deficient microvascular endothelium. J Cell Physiol. 2009, 220: 357-366. 10.1002/jcp.21772.PubMedCrossRef
22.
go back to reference Osada T, Gu YH, Kanazawa M, Tsubota Y, Hawkins BT, Spatz M, Milner R, Del Zoppo GJ: Interendothelial claudin-5 expression depends on cerebral endothelial cell-matrix adhesion by beta(1)-integrins. J Cereb Blood Flow Metab. 2001, 31: 1972-1985.CrossRef Osada T, Gu YH, Kanazawa M, Tsubota Y, Hawkins BT, Spatz M, Milner R, Del Zoppo GJ: Interendothelial claudin-5 expression depends on cerebral endothelial cell-matrix adhesion by beta(1)-integrins. J Cereb Blood Flow Metab. 2001, 31: 1972-1985.CrossRef
23.
go back to reference Urich E, Lazic SE, Molnos J, Wells I, Freskgard PO: Transcriptional profiling of human brain endothelial cells reveals key properties crucial for predictive in vitro blood–brain barrier models. PloS One. 2012, 7: e38149-10.1371/journal.pone.0038149.PubMedCentralPubMedCrossRef Urich E, Lazic SE, Molnos J, Wells I, Freskgard PO: Transcriptional profiling of human brain endothelial cells reveals key properties crucial for predictive in vitro blood–brain barrier models. PloS One. 2012, 7: e38149-10.1371/journal.pone.0038149.PubMedCentralPubMedCrossRef
24.
go back to reference Subileau E, Rezaie P, Davies H, Colyer F, Greenwood J, Male D, Romero I: Expression of chemokines and their receptors by human brain endothelium: implications for multiple sclerosis. J Neuropathol Exp Neurol. 2009, 68: 227-240. 10.1097/NEN.0b013e318197eca7.PubMedCrossRef Subileau E, Rezaie P, Davies H, Colyer F, Greenwood J, Male D, Romero I: Expression of chemokines and their receptors by human brain endothelium: implications for multiple sclerosis. J Neuropathol Exp Neurol. 2009, 68: 227-240. 10.1097/NEN.0b013e318197eca7.PubMedCrossRef
25.
go back to reference Pan W, Yu C, Hsuchou H, Khan RS, Kastin AJ: Cerebral microvascular IL15 is a novel mediator of TNF action. J Neurochem. 2009, 111: 819-827. 10.1111/j.1471-4159.2009.06371.x.PubMedCentralPubMedCrossRef Pan W, Yu C, Hsuchou H, Khan RS, Kastin AJ: Cerebral microvascular IL15 is a novel mediator of TNF action. J Neurochem. 2009, 111: 819-827. 10.1111/j.1471-4159.2009.06371.x.PubMedCentralPubMedCrossRef
Metadata
Title
Cytokine-induced changes in the gene expression profile of a human cerebral microvascular endothelial cell-line, hCMEC/D3
Authors
Miguel Alejandro Lopez-Ramirez
David Kingsley Male
Chunfang Wang
Basil Sharrack
Dongsheng Wu
Ignacio Andres Romero
Publication date
01-12-2013
Publisher
BioMed Central
Published in
Fluids and Barriers of the CNS / Issue 1/2013
Electronic ISSN: 2045-8118
DOI
https://doi.org/10.1186/2045-8118-10-27

Other articles of this Issue 1/2013

Fluids and Barriers of the CNS 1/2013 Go to the issue